Process for enriching basophils in a blood sample

Abstract

The present invention is based on the discovery that contrary to prior studies CD16 is expressed by basophils and as such provide methods and kits for enriching basophils in a blood sample.

1 . A method for enriching basophils in a blood sample, comprising
 (a) contacting the sample with an antibody cocktail which comprises antibodies that specifically bind to non-basophil cells in the sample and does not include any anti-CD16 antibodies,   (b) separating the non-basophil cells from the sample to obtain a blood sample enriched with basophils.   
   
   
       2 . The method of  claim 1 , wherein the antibody cocktail comprises at least one antibody which specifically binds to neutrophils, at least one antibody which specifically binds to natural killer cells, at least one antibody which specifically binds to monocytes, at least one antibody which specifically binds to dendritic cells, at least one antibody which specifically binds to T lymphocytes, at least one antibody which specifically binds to B lymphocytes, and at least one antibody which specifically binds to eosinophils. 
   
   
       3 . The method of  claim 1 , wherein the antibody cocktail further comprises at least one antibody which specifically binds to erythrocytes and at least one antibody which specifically binds to platelets. 
   
   
       4 . The method of  claim 1 , wherein the antibody cocktail comprises
 at least one antibody specific for CD14,   at least one antibody specific for CD24, and   at least one antibody specific for CD56.   
   
   
       5 . The method of  claim 1 , wherein the antibody cocktail comprises:
 at least one antibody specific for CD3,   at least one antibody specific for CD4,   at least one antibody specific for CD7,   at least one antibody specific for CD14,   at least one antibody specific for CD15,   at least one antibody specific for CD24,   at least one antibody specific for CD36,   at least one antibody specific for CD45RA,   at least one antibody specific for HLA-DR,   at least one antibody specific for CD56, and   at least one antibody specific for CD235a.   
   
   
       6 . The method of any one of  claim 1 , wherein the antibody cocktail comprises:
 at least one antibody specific for CD2,   at least one antibody specific for CD3,   at least one antibody specific for CD14,   at least one antibody specific for CD15,   at least one antibody specific for CD19,   at least one antibody specific for CD24,   at least one antibody specific for CD34,   at least one antibody specific for CD36,   at least one antibody specific for CD45RA,   at least one antibody specific for CD56, and   at least one antibody specific for CD235a.   
   
   
       7 . The method of  claim 1 , wherein the antibodies are monoclonal antibodies. 
   
   
       8 . The method of  claim 1 , wherein the antibodies are conjugated to magnetic beads. 
   
   
       9 . The method of  claim 8 , wherein the separating comprises applying a magnetic field to deplete the blood sample of non-basophil cells. 
   
   
       10 . The method of  claim 1 , wherein the antibodies are conjugated to biotin, and the separating comprises contacting the blood sample with anti-biotin antibodies conjugated to a magnetic bead and applying a magnetic field to deplete the sample of non-basophil cells. 
   
   
       11 . The method of  claim 1 , wherein the blood sample is peripheral blood, buffy coat, peripheral blood mononuclear cells (PBMC), or umbilical cord blood. 
   
   
       12 . The method of  claim 1 , further comprising before (a), the blood sample is pre-enriched in mononuclear cells by density gradient centrifugation, counter current centrifugal elutriation, erythrocytes lysis, Percoll gradients, or a combination thereof. 
   
   
       13 . The method of  claim 1 , further comprising after (b), evaluating the purity of the enriched basophils. 
   
   
       14 . The method of  claim 1 , wherein the anti-CD16 antibodies are specific for CD16B. 
   
   
       15 . A kit for enriching basophils in a blood sample, comprising antibodies that specifically bind to non-basophil cells but excluding any anti-CD16 antibody. 
   
   
       16 . The kit of  claim 15 , wherein the anti-CD16 antibody is specific for a CD16B isoform. 
   
   
       17 . The kit of  claim 15 , comprising at least one antibody specific for CD14, at least one antibody specific for CD24, and at least one antibody specific for CD56. 
   
   
       18 . The kit of  claim 15 , comprising at least one antibody specific for CD3, at least one antibody specific for CD4, at least one antibody specific for CD7, at least one antibody specific for CD14, at least one antibody specific for CD15, at least one antibody specific for CD24, at least one antibody specific for CD36, at least one antibody specific for CD45RA, at least one antibody specific for HLA-DR, at least one antibody specific for CD56, and at least one antibody specific for CD235a. 
   
   
       19 . The kit of  claim 15 , comprising at least one antibody specific for CD2, at least one antibody specific for CD3, at least one antibody specific for CD14, at least one antibody specific for CD15, at least one antibody specific for CD19, at least one antibody specific for CD24, at least one antibody specific for CD34, at least one antibody specific for CD36, at least one antibody specific for CD45RA, at least one antibody specific for CD56, and at least one antibody specific for CD235a. 
   
   
       20 . The kit of  claim 15 , wherein the antibodies are monoclonal antibodies. 
   
   
       21 . The kit of  claim 15 , wherein the antibodies are conjugated to magnetic beads. 
   
   
       22 . The kit of  claim 15 , wherein the antibodies are conjugated to biotin. 
   
   
       23 . The kit of  claim 15 , wherein the antibodies are physically separated or cocktailed to comprise two or more antibodies.